Reconstruction of skeletal defects represents a major clinical challenge with over 1 million surgical procedures performed each year. New strategies of regenerating bone are needed because of limitations with existing techniques. One new strategy is to create a composite graft in which autologous cells are seeded onto a porous, degradable scaffold. The scaffold supports the cells, structurally and biologically, allowing them to grow and secrete new extracellular matrix. Optimally tissue growth occurs concurrent with scaffold degradation. The degree of new bone formation is, however, material dependent and not predictable. We therefore seek to establish material chemistry parameters that could optimize bone cell function. In pursuit of this goal, we have developed: (1) in vitro culture methods in which human bone marrow stromal cells (BMSCs) are expanded; (2) polymer processing techniques to reproducibly fabricate highly porous 3D poly(lactic-co-glycolic) scaffolds, which have been successfully used to engineer a number of tissues including bone; (3) materials science design strategies which enable us to biomimetically modify both the internal microenvironment of a scaffold and the scaffold surface; and (4) a critical size cranial defect model in an immunocompromised mouse which has shown that the human BMSCs are capable of forming new bone in an animal model. The global hypothesis of the proposed research is that the extracellular microenvironment provided by the scaffold modulates the ability of human BMSCs to differentiate toward an osteoblast phenotype, and therefore controls biomineralization and structural integrity of regenerated bone. Results from our and other laboratories support this hypothesis, which is tested by synthesizing a series of model biomimetic materials. First, we synthesize environmentally responsive or "smart" scaffolds that buffer the microenvironment upon scaffold degradation. Second, we synthesize scaffolds with a surface that self-mineralizes into a biological apatite. Third, we use functionally-graded scaffolds in which mineralization is spatially controlled. The rationale for each of these 3 biomimetic strategies lies in the way nature has designed the skeleton. The skeletal system is able to perform its functions using a minimum amount of mass because biology has utilized design approaches, which include the ability to adapt to environmental cues (i.e. "smartness"), a hierarchical organization consisting of elegant mineral synthesis, and an organization that is optimized for physiological function by having gradients in composition and structure. In the proposed studies, we aim to exploit aspects of each of these 3 biomimetic strategies in an effort to create biomaterials that will modulate biological response in a controlled manner.